Effective population size and evolutionary dynamics in outbred laboratory populations of Drosophila

Census population size, sex-ratio and female reproductive success were monitored in 10 laboratory populations of Drosophila melanogaster selected for different ages of reproduction. With this demographic information, we estimated eigenvalue, variance and probability of allele loss effective population sizes. We conclude that estimates of effective size based on gene-frequency change at a few loci are biased downwards. We analysed the relative roles of selection and genetic drift in maintaining genetic variation in laboratory populations of Drosophila. We suggest that rare, favourable genetic variants in our laboratory populations have a high chance of being lost if their fitness effect is weak, e.g. 1% or less. However, if the fitness effect of this variation is 10% or greater, these rare variants are likely to increase to high frequency. The demographic information developed in this study suggests that some of our laboratory populations harbour more genetic variation than expected. One explanation for this finding is that part of the genetic variation in these outbred laboratory Drosophila populations may be maintained by some form of balancing selection. We suggest that, unlike bacteria, medium-term adaptation of laboratory populations of fruit flies is not primarily driven by new mutations, but rather by changes in the frequency of preexisting alleles.     

Variation of body size within and between wild populations of Drosophila buzzatii

Four populations of the cactophilous species D. buzzatii have been compared with respect to the phenotypic variation of thorax and wing length of wild versus laboratory reared flies. Three of the strains were intercrossed to provide parent, F₁ and F₂ comparisons as a test of co-adaptation. The genetic contribution to phenotypic variation of laboratory reared flies was estimated from the correlation between sibs derived from random pair mating and reared individually in separate cultures. The average natural temperature during development was estimated from the relations between the wing/thorax ratio and temperature in laboratory tests.The variance of thorax and wing length of wild flies was several times greater than that of laboratory reared flies and the increase was attributed primarily to variation in larval food supply although temperature fluctuation is also important. There was no evidence of heterosis or F₂ break-down in the crosses. For two of the populations the heritability of thorax length was high, 60–70%, and substantially lower for the third. The average temperature estimated from the wing/thorax/temperature relationship differed between sites. The reduction of body size below the potential maximum averaged 30% for two and 20% for the other population, with a wide spread about these values. The evidence is discussed in relation to assessing the nature of ecological variation by comparing the variation of morphological traits in wild and laboratory reared flies.     

SNP allele frequency estimation in DNA pools and variance components analysis

The estimation of single nucleotide polymorphism (SNP) allele frequency in pooled DNA samples has been proposed as a cost-effective approach to whole genome association studies. However, the key issue is the allele frequency window in which a genotyping method operates and provides a statistically reliable answer. We assessed the homogeneous mass extend assay and estimated the variance associated with each experimental stage. We report that a relationship between estimated allele frequency and variance might exist, suggesting that high statistical power can be retained at low, as well as high, allele frequencies. Assuming this relationship, the formation of subpools consisting of 100 samples retains an effective sample size greater than 70% of the true sample size, with a savings of 11-fold the cost of an individual genotyping study, regardless of allele frequency.     

Seasonal changes in the body size of two rotifer species living in activated sludge follow the Temperature‐Size Rule

Temperature‐Size Rule (TSR) is a phenotypic body size response of ectotherms to changing temperature. It is known from the laboratory studies, but seasonal patterns in the field were not studied so far. We examined the body size changes in time of rotifers inhabiting activated sludge. We hypothesize that temperature is the most influencing parameter in sludge environment, leading sludge rotifers to seasonally change their body size according to TSR, and that oxygen content also induces the size response. The presence of TSR in Lecane inermis rotifer was tested in a laboratory study with two temperature and two food‐type treatments. The effect of interaction between temperature and food was significant; L. inermis followed TSR in one food type only. The seasonal variability in the body sizes of the rotifers L. inermis and Cephalodella gracilis was estimated by monthly sampling and analyzed by multiple regression, in relation to the sludge parameters selected as the most influential by multivariate analysis, and predicted to alter rotifer body size (temperature and oxygen). L. inermis varied significantly in size throughout the year, and this variability is explained by temperature as predicted by the TSR, but not by oxygen availability. C. gracilis also varied in size, though this variability was explained by both temperature and oxygen. We suggest that sludge age acts as a mortality factor in activated sludge. It may have a seasonal effect on the body size of L. inermis and modify a possible effect of oxygen. Activated sludge habitat is driven by both biological processes and human regulation, yet its resident organisms follow general evolutionary rule as they do in other biological systems. The interspecific response patterns differ, revealing the importance of taking species‐specific properties into account. Our findings are applicable to sludge properties enhancement through optimizing the conditions for its biological component.     

In situ versus laboratory estimations of length–weight regression and growth rate of Daphnia magna (Branchiopoda, Anomopoda) from an aerated waste stabilization pond

The present paper questions the adequacy of using length–weight regressions and growth rates calculated in the laboratory under constant physico-chemical and food conditions for the estimation of biomass and secondary production of animals living in a variable environment from the physico-chemistry and food availability point of view. Length–weight regressions (LWR) and growth rate of Daphnia magna were determined in situat five key periods of the year. In parallel, LWR and growth rate were determined in laboratory incubators at temperature adjusted to the mean temperature measured during the in situexperiments. LWR estimated from pond daphnids collected during the in situ experiments were, on the whole, not significantly different from LWR established during laboratory experiments, indicating that the food availability was globally similar in the laboratory and in situexperiments, even though food items were substantially different between the experiments. In situ algal biomass was indeed low compared to the algal biomass in laboratory experiments, but high biomasses of bacteria, protozoa and detritus were available for daphnid feeding in the tubes incubated in situ. Growth rate of D. magnawas monitored in situusing 50-ml tubes closed with Nylon net (mesh size = 80 m) and in the laboratory using 50-ml glass flasks. The physico-chemical, bacteriological and algological variables were checked to be similar in the tubes and in the pond. Growth rates varied according to the size of the animal and according to the water temperature. The maximum growth rates were observed for juveniles at 20.2 °C. Growth rates were also determined in the laboratory at temperature corresponding to the mean temperature recorded in the pond during the in situ growth experiments. Differences between in situ and laboratory body length–growth rate regressions (LgR) were observed for the experiments conducted at 15.6 °C and 23.6 °C. Due to differences in LWR and LgR between in situ and laboratory experiments, biomass and daily production estimated from laboratory cultures were found to be significantly, but not severely, higher than biomass and daily production estimated on the basis of in situ experiments. It has been, therefore, concluded that, when the constraints linked to the realization of in situ growth experiments are too strong, the laboratory approach is fully justified.     

Demographic Genetics of Brown Trout (Salmo Trutta) and Estimation of Effective Population Size from Temporal Change of Allele Frequencies

We studied temporal allele frequency shifts over 15 years and estimated the genetically effective size of four natural populations of brown trout (Salmo trutta L.) on the basis of the variation at 14 polymorphic allozyme loci. The allele frequency differences between consecutive cohorts were significant in all four populations. There were no indications of natural selection, and we conclude that random genetic drift is the most likely cause of temporal allele frequency shifts at the loci examined. Effective population sizes were estimated from observed allele frequency shifts among cohorts, taking into consideration the demographic characteristics of each population. The estimated effective sizes of the four populations range from 52 to 480 individuals, and we conclude that the effective size of natural brown trout populations may differ considerably among lakes that are similar in size and other apparent characteristics. In spite of their different effective sizes all four populations have similar levels of genetic variation (average heterozygosity) indicating that excessive loss of genetic variability has been retarded, most likely because of gene flow among neighboring populations.     

Clutch size decisions of a gregarious parasitoid under laboratory and field conditions

Under field conditions, insect parasitoids probably experience lower rates of host encounter and life expectancy than under optimal conditions in laboratory studies. We examined the clutch size response of Mastrus ridibundus, a gregarious idiobiont parasitoid of codling moth, Cydia pomonella, cocoons, to variation in both host encounter rate and life expectancy as possible explanatory variables in a comparison of brood size in the field and laboratory. Under laboratory conditions, mean clutch size (number of eggs laid) declined from 5.8 to 3.4 as host encounter rate increased from one to eight cocoons per day. In contrast, when we reduced life expectancy by withholding honey as a food source, females did not adjust clutch size. Mean brood size (number of progeny surviving to pupation) of females foraging in walnut orchards (3.9) was significantly greater than that under laboratory conditions with excess hosts (3.1). Brood size also increased with host size in the field, but not under laboratory conditions. Brood size fitness curves were derived using both host-finding ability in the field and lifetime fecundity under laboratory conditions as indices of female fitness. Host-finding ability increased exponentially with body size, generating an estimated Lack brood size of 4.3, but lifetime fecundity increased linearly with body size, giving a Lack brood size estimate of 5.5. Under field conditions, female M. ridibundus produced brood sizes that closely approximated the Lack brood size estimated from host-finding ability, but that were significantly smaller than that estimated from lifetime fecundity. These observations suggest that, in contrast to lifetime fecundity measures from the laboratory, host-finding ability in the field provides a more accurate estimate of lifetime reproductive success for parasitoids with a low expectation of future reproduction. Copyright 2003 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.      

Evaluation of DNA pooling for the estimation of microsatellite allele frequencies: a case study using striped bass (Morone saxatilis)

Using striped bass (Morone saxatilis) and six multiplexed microsatellite markers, we evaluated procedures for estimating allele frequencies by pooling DNA from multiple individuals, a method suggested as cost-effective relative to individual genotyping. Using moment-based estimators, we estimated allele frequencies in experimental DNA pools and found that the three primary laboratory steps, DNA quantitation and pooling, PCR amplification, and electrophoresis, accounted for 23, 48, and 29%, respectively, of the technical variance of estimates in pools containing DNA from 2-24 individuals. Exact allele-frequency estimates could be made for pools of sizes 2-8, depending on the locus, by using an integer-valued estimator. Larger pools of size 12 and 24 tended to yield biased estimates; however, replicates of these estimates detected allele frequency differences among pools with different allelic compositions. We also derive an unbiased estimator of Hardy-Weinberg disequilibrium coefficients that uses multiple DNA pools and analyze the cost-efficiency of DNA pooling. DNA pooling yields the most potential cost savings when a large number of loci are employed using a large number of individuals, a situation becoming increasingly common as microsatellite loci are developed in increasing numbers of taxa.     

The relative quantities and catalytic activities of enzymes produced by alleles at the alcohol dehydrogenase locus in Drosophila melanogaster

We have examined the kinetic properties of enzymes produced by the electrophoretically fast (F) and slow (S) alleles at the alcohol dehydrogenase locus in a polymorphic laboratory population of Drosophila melanogaster. The product of the F allele has approximately twice the specific activity of the product of the S allele. We have estimated four Michaelis constants (K _ethanol, K _NAD, K′_ethanol, and K′_NAD) and have found no significant difference between the major (ADH-5) isozyme produced by homozygotes for the two alleles. The relative amounts of enzyme produced by the homozygotes were estimated by the method of Laurell (1966), and again no significant differences were found. It appears that the difference in specific activities can be explained solely in terms of relative catalytic efficiency. In a series of laboratory stocks, those fixed for the F allele tend to produce more enzyme than those fixed for the S allele. These observations do not support the view that the alleles are selectively equivalent.     

Sample size considerations in genetic polymorphism studies.

OBJECTIVES: Molecular studies for genetic polymorphisms are being carried out for a number of different applications, such as genetic disorders in different populations, pharmacogenomics, genetic identification of ethnic groups for forensic and legal applications, genetic identification of breed/stock in animals and plants for commercial applications and conservation of germ plasm. In this paper, for a random sampling scheme, we address two questions: (A) What should be the minimum size of the sample so that, with a prespecified probability, all alleles at a given locus (or haplotypes at a given set of loci) are detected? (B) What should be the sample size so that the allele frequency distribution at a given locus (or haplotype frequency distribution at a given set of loci) is estimated reliably within permissible error limits? METHODS: We have used combinatorial probabilistic arguments and Monte Carlo simulations to answer these questions. RESULTS: We found that the minimum sample size required in case A depends mainly on the prespecified probability of detecting all alleles, while in case B, it varies greatly depending on the permissible error in estimation (which will vary with the application). We have obtained the minimum sample sizes for different degrees of polymorphism at a locus under high stringency, as well as a relaxed level of permissible error. We present a detailed sampling procedure for estimating allele frequencies at a given locus, which will be of use in practical applications. CONCLUSION: Since the sample size required for reliable estimation of allele frequency distribution increases with the number of alleles at the locus, there is a strong case for using biallelic markers (like single nucleotide polymorphisms) when the available sample size is about 800 or less.     

DOES MATE LIMITATION IN SELF‐INCOMPATIBLE SPECIES PROMOTE THE EVOLUTION OF SELFING? THE CASE OF LEAVENWORTHIA ALABAMICA

Genetic diversity at the S‐locus controlling self‐incompatibility (SI) is often high because of negative frequency‐dependent selection. In species with highly patchy spatial distributions, genetic drift can overwhelm balancing selection and cause stochastic loss of S‐alleles. Natural selection may favor the breakdown of SI in populations with few S‐alleles because low S‐allele diversity constrains the seed production of self‐incompatible plants. We estimated S‐allele diversity, effective population sizes, and migration rates in Leavenworthia alabamica, a self‐incompatible mustard species restricted to discrete habitat patches in rocky glades. Patterns of polymorphism were investigated at the S‐locus and 15 neutral microsatellites in three large and three small populations with 100‐fold variation in glade size. Populations on larger glades maintained more S‐alleles, but all populations were estimated to harbor at least 20 S‐alleles, and mate availabilities typically exceeded 0.80, which is consistent with little mate limitation in nature. Estimates of the effective size (N_e) in each population ranged from 600 to 1600, and estimated rates of migration (m) ranged from 3 × 10⁻⁴ to nearly 1 × 10⁻³. According to theoretical models, there is limited opportunity for genetic drift to reduce S‐allele diversity in populations with these attributes. Although pollinators or resources limit seed production in small glades, limited S‐allele diversity does not appear to be a factor promoting the incipient breakdown of SI in populations of this species that were studied.     

TEMPORAL CHANGE OF MITOCHONDRIAL DNA HAPLOTYPE FREQUENCIES AND FEMALE EFFECTIVE SIZE IN A BROWN TROUT (SALMO TRUTTA) POPULATION

We report data on genetic drift of mitochondrial DNA (mtDNA) haplotypes in a natural brown trout (Salmo trutta) population in Sweden. Large temporal frequency shifts were observed over the 14 consecutive year classes studied. The observed rate of drift was used to estimate the effective size of the population. This effective size applies to the female segment of the population as mtDNA is maternally inherited. The magnitude of mtDNA haplotype frequency change is compared with the corresponding allele frequency changes at 14 allozyme loci in the same population. The female effective size is estimated as 58, which is approximately half the effective size of 97 for the total population (both sexes) previously obtained from the shifts of allozyme allele frequencies.     

Laboratory energetics of larvae of Sericostoma personatum (Trichoptera)

The energetics of the shredder Sericostoma personatum Spence were studied in the laboratory at 3,8, 13 and 18°C using conditioned leaves of Fagus sylvatica L. as food. Ingestion (I), growth (G) and respiration (R) were dependent on larval size and temperature. Estimated assimilation (G + R) varied between 4.2–5.7%. Gross growth efficiency (G/I × 100) varied between 0.4–3.4% and was dependent on temperature. Growth rates estimated from field and laboratory data were in the same order of magnitude.
The low assimilation is discussed in relation to the life cycle pattern of the species and its ecological role in the stream community.     

A large‐scale latitudinal pattern of life‐history traits in a strictly univoltine damselfly

1. Variation in thermal conditions and season length along latitudinal gradients affect body size‐related traits over different life stages. Selection is expected to optimise these size traits in response to the costs and benefits. 2. Egg, hatchling, larval and adult size in males and females were estimated along a latitudinal gradient of 2730 km across Europe in the univoltine damselfly Lestes sponsa, using a combination of field‐collection and laboratory‐rearing experiments. In the laboratory, individuals were grown in temperatures and photoperiod simulating those at the latitude of origin, and in common‐garden conditions. 3. The size of adults sampled in nature was negatively correlated with latitude. In all populations the females were larger than the males. Results from simulated and common‐garden rearing experiments supported this pattern of size difference across latitudes and between sexes, suggesting a genetic component for the latitudinal size trend and female‐biased size dimorphism. In contrast, hatchling size showed a positive relationship with latitude, but egg size, although differing between latitudes, showed no such relationship. 4. The results support a converse Bergmann cline, i.e. a negative body size cline towards the north. This negative cline in body size is probably driven by progressively stronger seasonal time and temperature constraints towards the higher latitudes and by the obligate univoltine life cycle of L. sponsa. As egg size showed no relationship with latitude, other environmental factors besides temperature, such as desiccation risk, probably affect this trait.     

Estimating genetic drift and effective population size from temporal shifts in dominant gene marker frequencies

Measurement of temporal change in allele frequencies represents an indirect method for estimating the genetically effective size of populations. When allele frequencies are estimated for gene markers that display dominant gene expression, such as, e.g. random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) markers, the estimates can be seriously biased. We quantify bias for previous allele frequency estimators and present a new expression that is generally less biased and provides a more precise assessment of temporal allele frequency change. We further develop an estimator for effective population size that is appropriate when dealing with dominant gene markers. Comparison with estimates based on codominantly expressed genes, such as allozymes or microsatellites, indicates that about twice as many loci or sampled individuals are required when using dominant markers to achieve the same precision.     

The power and efficiency of brood incubation in queenless microcolonies of bumble bees (Bombus terrestris L.)

1. Ground‐nesting colonies of bumble bees incubate their brood at > 30 °C if floral forage provides sufficient energy and the thermogenic power of the colony can counteract cool soil conditions. To explore the basis of incubation, the thermogenic power and sugar consumption of orphaned nests of bumble bee workers (microcolonies) were investigated under laboratory conditions. 2. This study tested experimentally the effect of variation in worker number (ranging from four to 12 adults) on a microcolony's capacity to regulate brood temperature and recover from acute cold exposure. Microcolonies were provided with ad libitum sugar syrup and minimal insulation and maintained at an ambient temperature of c. 25 °C. Energy conversion efficiency was estimated by comparing sugar consumption with the power required for artificial incubation. The joint energetics of foraging and incubation were modelled in wild colonies to explore the effect of colony size and landscape quality on thermoregulation. 3. The results showed that all sizes of microcolonies regulated brood temperature at c. 31 °C under laboratory conditions, which required 96 mW of thermogenic power. It was estimated that individual workers of B. terrestris generated an incubatory power of 35 mW. The smallest microcolonies had the highest conversion efficiency (57%), apparently because few workers were required for incubation. 4. Modelling indicated that small microcolonies of three to seven adult workers have the capacity for normal brood incubation in the wild, but that the minimum viable colony size increases as floral forage becomes poorer or more distant. 5. These preliminary findings suggest the feasibility of identifying the minimum conditions (forage quality, soil temperature, and colony size) necessary for brood incubation by queenright colonies in the wild.     

Long-term genetic stability and population dynamics of laboratory strains of Schistosoma mansoni

Measures of genetic differentiation between populations are useful tools for understanding the long-term dynamics of parasite communities. We followed the allele frequencies of microsatellite markers in samples taken over a period of 16 yr from the Case Western Reserve University-Naval Medical Research Institute (CWRU-NMRI) laboratory strain of Schistosoma mansoni. DNA was isolated from pooled samples of adults, eggs, or cercariae collected at 46 time points and genotyped for 14 tri- or tetranucleotide microsatellite markers. For comparison, 2 S. mansoni reference strains (Biomedical Research Institute-NMRI, which has a common origin with the CWRU line, and PR-1) were analyzed over shorter periods of time. We observed that the long-term allele frequencies are generally stable in large laboratory populations of this parasite, and a high degree of similarity was observed between the allele frequencies of consecutive samples from different developmental stages. The CWRU strain, however, showed 2 periods of marked deviation from stability as demonstrated using genetic differentiation measures. The first period corresponds to an admixture event with the BRI strain in which a new equilibrium was established as the "migrants" became blended into the existing CWRU population, consistent with 23% admixture from BRI. The second corresponds to a period of genetic drift when the CWRU population size was greatly reduced with an accompanying loss in genetic diversity. Having demonstrated the utility of pooled samples for the genetic analysis of population dynamics in laboratory strains of schistosomes, this approach will be useful for analyzing field samples to determine the impact of schistosomiasis control programs on parasite population structure. Accounting only for the intensity or prevalence of parasite populations may fail to register significant changes in population structure that could have implications for resistance, morbidity, and the design of control measures.     

Hatching plasticity in the tropical gastropod Nerita scabricosta

Hatching plasticity has been documented in diverse terrestrial and freshwater taxa, but in few marine invertebrates. Anecdotal observations over the last 80 years have suggested that intertidal neritid snails may produce encapsulated embryos able to significantly delay hatching. The cause for delays and the cues that trigger hatching are unknown, but temperature, salinity, and wave action have been suggested to play a role. We followed individual egg capsules of Nerita scabricosta in 16 tide pools to document the variation in natural time to hatching and to determine if large delays in hatching occur in the field. Hatching occurred after about 30 d and varied significantly among tide pools in the field. Average time to hatching in each pool was not correlated with presence of potential predators, temperature, salinity, or pool size. We also compared hatching time between egg capsules in the field to those kept in the laboratory at a constant temperature in motionless water, and to those kept in the laboratory with sudden daily water motion and temperature changes. There was no significant difference in the hatching rate between the two laboratory treatments, but capsules took, on average, twice as long to hatch in the laboratory as in the field. Observations of developing embryos showed that embryos in the field develop slowly and continuously until hatching, but embryos in the laboratory reach the hatching stage during the first month of development and remain in stasis after that. Instances of hatching plasticity in benthic marine invertebrates, like the one in N. scabricosta, could greatly enhance our ability to investigate the costs and benefits of benthic versus planktonic development, a long‐standing area of interest for invertebrate larval biologists.     

Effective population size associated with self-fertilization: lessons from temporal changes in allele frequencies in the selfing annual Medicago truncatula

Despite its significance in evolutionary and conservation biology, few estimates of effective population size (N(e)) are available in plant species. Self-fertilization is expected to affect N(e), through both its effect on homozygosity and population dynamics. Here, we estimated N(e) using temporal variation in allele frequencies for two contrasted populations of the selfing annual Medicago truncatula: a large and continuous population and a subdivided population. Estimated N(e) values were around 5-10% of the population census size suggesting that other factors than selfing must contribute to variation in allele frequencies. Further comparisons between monolocus allelic variation and changes in the multilocus genotypic composition of the populations show that the local dynamics of inbred lines can play an important role in the fluctuations of allele frequencies. Finally, comparing N(e) estimates and levels of genetic variation suggest that H(e) is a poor estimator of the contemporaneous variance effective population size.     

Importance of a pilot study for non-invasive genetic sampling: genotyping errors and population size estimation in red deer

Population size information is critical for managing endangered or harvested populations. Population size can now be estimated from non-invasive genetic sampling. However, pitfalls remain such as genotyping errors (allele dropout and false alleles at microsatellite loci). To evaluate the feasibility of non-invasive sampling (e.g., for population size estimation), a pilot study is required. Here, we present a pilot study consisting of (i) a genetic step to test loci amplification and to estimate allele frequencies and genotyping error rates when using faecal DNA, and (ii) a simulation step to quantify and minimise the effects of errors on estimates of population size. The pilot study was conducted on a population of red deer in a fenced natural area of 5440 ha, in France. Twelve microsatellite loci were tested for amplification and genotyping errors. The genotyping error rates for microsatellite loci were 0–0.83 (mean=0.2) for allele dropout rates and 0–0.14 (mean=0.02) for false allele rates, comparable to rates encountered in other non-invasive studies. Simulation results suggest we must conduct 6 PCR amplifications per sample (per locus) to achieve approximately 97% correct genotypes. The 3% error rate appears to have little influence on the accuracy and precision of population size estimation. This paper illustrates the importance of conducting a pilot study (including genotyping and simulations) when using non-invasive sampling to study threatened or managed populations.